Skip to main content
PMC home page
Applied and Environmental Microbiology logoLink to Applied and Environmental Microbiology
. 1997 May;63(5):2062–2066. doi: 10.1128/aem.63.5.2062-2066.1997

Molecular analysis of the Rhodococcus sp. strain H1 her gene and characterization of its product, a heroin esterase, expressed in Escherichia coli.

D A Rathbone 1, P J Holt 1, C R Lowe 1, N C Bruce 1
PMCID: PMC168495  PMID: 9143135

Abstract

The structural gene for heroin esterase was cloned from Rhodococcus sp. strain H1 and expressed in Escherichia coli BL21(DE3). The purified enzyme was found to be a tetramer with an M(r) of 137,000 and an apparent K(m) of 0.88 mM for 6-acetylmorphine. The G-x-S-x-G motif was observed in the deduced amino acid sequence, suggesting that the enzyme is a serin esterase.

Full Text

The Full Text of this article is available as a PDF (317.2 KB).

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Bruce N. C., Caswell D. A., French C. E., Hailes A. M., Long M. T., Willey D. L. Towards engineering pathways for the synthesis of analgesics and antitussives. Ann N Y Acad Sci. 1994 May 2;721:85–99. doi: 10.1111/j.1749-6632.1994.tb47380.x. [DOI] [PubMed] [Google Scholar]
  2. Bruce N. C., Willey D. L., Coulson A. F., Jeffery J. Bacterial morphine dehydrogenase further defines a distinct superfamily of oxidoreductases with diverse functional activities. Biochem J. 1994 May 1;299(Pt 3):805–811. doi: 10.1042/bj2990805. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Bruce N. C., Wilmot C. J., Jordan K. N., Stephens L. D., Lowe C. R. Microbial degradation of the morphine alkaloids. Purification and characterization of morphine dehydrogenase from Pseudomonas putida M10. Biochem J. 1991 Mar 15;274(Pt 3):875–880. doi: 10.1042/bj2740875. [DOI] [PMC free article] [PubMed] [Google Scholar]
  4. Bruce N. C., Wilmot C. J., Jordan K. N., Trebilcock A. E., Gray Stephens L. D., Lowe C. R. Microbial degradation of the morphine alkaloids: identification of morphine as an intermediate in the metabolism of morphine by Pseudomonas putida M10. Arch Microbiol. 1990;154(5):465–470. doi: 10.1007/BF00245229. [DOI] [PubMed] [Google Scholar]
  5. Cameron G. W., Jordan K. N., Holt P. J., Baker P. B., Lowe C. R., Bruce N. C. Identification of a heroin esterase in Rhodococcus sp. strain H1. Appl Environ Microbiol. 1994 Oct;60(10):3881–3883. doi: 10.1128/aem.60.10.3881-3883.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  6. Feller G., Thiry M., Gerday C. Nucleotide sequence of the lipase gene lip2 from the antarctic psychrotroph Moraxella TA144 and site-specific mutagenesis of the conserved serine and histidine residues. DNA Cell Biol. 1991 Jun;10(5):381–388. doi: 10.1089/dna.1991.10.381. [DOI] [PubMed] [Google Scholar]
  7. Hart S., Kirby R., Woods D. R. Structure of a Rhodococcus gene encoding pigment production in Escherichia coli. J Gen Microbiol. 1990 Jul;136(7):1357–1363. doi: 10.1099/00221287-136-7-1357. [DOI] [PubMed] [Google Scholar]
  8. Holt P. J., Bruce N. C., Lowe C. R. Bioluminescent assay for heroin and its metabolites. Anal Chem. 1996 Jun 1;68(11):1877–1882. doi: 10.1021/ac951207r. [DOI] [PubMed] [Google Scholar]
  9. Holt P. J., Stephens L. D., Bruce N. C., Lowe C. R. An amperometric opiate assay. Biosens Bioelectron. 1995 Summer;10(6-7):517–526. doi: 10.1016/0956-5663(95)96927-q. [DOI] [PubMed] [Google Scholar]
  10. Raibaud A., Zalacain M., Holt T. G., Tizard R., Thompson C. J. Nucleotide sequence analysis reveals linked N-acetyl hydrolase, thioesterase, transport, and regulatory genes encoded by the bialaphos biosynthetic gene cluster of Streptomyces hygroscopicus. J Bacteriol. 1991 Jul;173(14):4454–4463. doi: 10.1128/jb.173.14.4454-4463.1991. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Rathbone D. A., Holt P. J., Bruce N. C., Lowe C. R. The use of recombinant DNA technology in the design of a highly specific heroin sensor. Ann N Y Acad Sci. 1996 May 15;782:534–543. doi: 10.1111/j.1749-6632.1996.tb40591.x. [DOI] [PubMed] [Google Scholar]
  12. Rathbone D. A., Holt P. J., Lowe C. R., Bruce N. C. The use of a novel recombinant heroin esterase in the development of an illicit drugs biosensor. Ann N Y Acad Sci. 1996 Oct 12;799:90–96. doi: 10.1111/j.1749-6632.1996.tb33184.x. [DOI] [PubMed] [Google Scholar]
  13. Salvi S., Trinei M., Lanfaloni L., Pon C. L. Cloning and characterization of the gene encoding an esterase from Spirulina platensis. Mol Gen Genet. 1994 Apr;243(1):124–126. doi: 10.1007/BF00283885. [DOI] [PubMed] [Google Scholar]
  14. Schirmer A., Jendrossek D. Molecular characterization of the extracellular poly(3-hydroxyoctanoic acid) [P(3HO)] depolymerase gene of Pseudomonas fluorescens GK13 and of its gene product. J Bacteriol. 1994 Nov;176(22):7065–7073. doi: 10.1128/jb.176.22.7065-7073.1994. [DOI] [PMC free article] [PubMed] [Google Scholar]
  15. Schwartz D., Alijah R., Nussbaumer B., Pelzer S., Wohlleben W. The peptide synthetase gene phsA from Streptomyces viridochromogenes is not juxtaposed with other genes involved in nonribosomal biosynthesis of peptides. Appl Environ Microbiol. 1996 Feb;62(2):570–577. doi: 10.1128/aem.62.2.570-577.1996. [DOI] [PMC free article] [PubMed] [Google Scholar]
  16. Sobek H., Görisch H. Purification and characterization of a heat-stable esterase from the thermoacidophilic archaebacterium Sulfolobus acidocaldarius. Biochem J. 1988 Mar 1;250(2):453–458. doi: 10.1042/bj2500453. [DOI] [PMC free article] [PubMed] [Google Scholar]
  17. Studier F. W., Moffatt B. A. Use of bacteriophage T7 RNA polymerase to direct selective high-level expression of cloned genes. J Mol Biol. 1986 May 5;189(1):113–130. doi: 10.1016/0022-2836(86)90385-2. [DOI] [PubMed] [Google Scholar]
  18. Walker C. H., Mackness M. I. Esterases: problems of identification and classification. Biochem Pharmacol. 1983 Nov 15;32(22):3265–3269. doi: 10.1016/0006-2952(83)90349-0. [DOI] [PubMed] [Google Scholar]
  19. Willey D. L., Caswell D. A., Lowe C. R., Bruce N. C. Nucleotide sequence and over-expression of morphine dehydrogenase, a plasmid-encoded gene from Pseudomonas putida M10. Biochem J. 1993 Mar 1;290(Pt 2):539–544. doi: 10.1042/bj2900539. [DOI] [PMC free article] [PubMed] [Google Scholar]
  20. Yanisch-Perron C., Vieira J., Messing J. Improved M13 phage cloning vectors and host strains: nucleotide sequences of the M13mp18 and pUC19 vectors. Gene. 1985;33(1):103–119. doi: 10.1016/0378-1119(85)90120-9. [DOI] [PubMed] [Google Scholar]

Articles from Applied and Environmental Microbiology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES